There are currently no licensed vaccines against Clostridium perfringens which causes necrotic enteritis in poultry. Chitosan nanoparticles were formulated with native (CN) or toxoids (CT) of extracellular proteins (ECP) of C. perfringens, both surface-tagged with Salmonella flagellar proteins. In a pH stability assay, CN and CT nanoparticles released 6% and 0% of their protein at 8.0 pH. In a protein release assay, CN and CT nanoparticles released 16% and 10% of their protein respectively at 7.4 pH after 24 h. CN and CT nanoparticles incubated at 100 μg/mL PBS with Chicken RBCs released 1% and 0% hemoglobin respectively. Ninety broilers were randomly assigned to treatments; sham-vaccinated (Control), CN-vaccinated (CN), and CT-vaccinated (CT). Each bird was orally gavaged with 50 μg vaccine in 0.5 mL PBS or 0.5 mL PBS only on d 0, 3, 7 and 14 of age. At 21 d of age, the CN group had higher anti-ECP IgA than control (P < 0.05). At 21 d of age, the CN and CT group had higher anti-ECP IgA than control (P < 0.05). At 17 d of age, the CN group had higher anti-flagellar IgG than control (P < 0.05). At 10 d of age, the CN group had higher anti-flagellar IgA than control (P < 0.05). Splenic T cells from chickens in the CN and CT group ex-vivo stimulated with 0.05 mg/mL ECP, had higher proliferation control (P < 0.05, P < 0.01 respectively). Splenic T cells from chickens in the CN and CT groups ex-vivo stimulated with 0.1 mg/mL ECP had proliferation than control (P < 0.05). Pooled serum from 17 d of age CN and CT-vaccinated birds partially neutralized toxins in 50 μg of ECP (P < 0.05). Pooled serum from 28 d of age CN-vaccinated birds also partially neutralized toxins in 50 μg of ECP. The result from this study indicates the potential for chitosan loaded with Clostridium perfringens extracellular proteins to be applied to necrotic enteritis challenge studies.

中文翻译：

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壳聚糖纳米颗粒的体外表征和免疫原性与坏死性肠炎相关的产气荚膜梭状芽胞杆菌田间菌株中负载了天然和灭活的细胞外蛋白。

目前尚无针对导致产家坏死性肠炎的产气荚膜梭状芽孢杆菌的许可疫苗。用产气荚膜梭菌胞外蛋白（ECP）的天然（CN）或类毒素（CT）配制壳聚糖纳米颗粒，二者均在表面标记有沙门氏菌鞭毛蛋白。在pH稳定性测定中，CN和CT纳米颗粒在8.0 pH下释放了6％和0％的蛋白质。在蛋白质释放测定中，CN和CT纳米粒子在24小时后在7.4 pH下分别释放其蛋白质的16％和10％。用鸡RBC在100μg/ mL PBS中孵育的CN和CT纳米颗粒分别释放1％和0％的血红蛋白。九十只肉鸡被随机分配进行治疗；假接种（对照），CN接种（CN）和CT接种（CT）。仅在第0、3、3天，每只鸡口服0.5μgPBS或0.5 mL PBS中的50μg疫苗 7岁和14岁。在21岁时，CN组的抗ECP IgA水平高于对照组（P <0.05）。在21 d时，CN和CT组的抗ECP IgA值高于对照组（P <0.05）。在17岁时，CN组的抗鞭毛IgG含量高于对照组（P <0.05）。在10 d龄时，CN组的抗鞭毛IgA水平高于对照组（P <0.05）。CN和CT组的鸡脾脏T细胞离体刺激0.05 mg / mL ECP，具有较高的增殖控制（分别为P <0.05，P <0.01）。CN和CT组的鸡脾脏T细胞在0.1 mg / mL ECP刺激下离体增殖（P <0.05）。来自17天龄的CN和CT疫苗接种的禽类的合并血清部分中和了50μgECP中的毒素（P <0.05）。来自28天龄经CN疫苗接种的禽类的合并血清也部分中和了50μgECP中的毒素。这项研究的结果表明，载有产气荚膜梭状芽胞杆菌胞外蛋白的壳聚糖有可能用于坏死性肠炎攻毒研究。